The RSV F glycoprotein is a key target for vaccine-induced neutralizing antibody. Our hypothesis is that understanding the mechanism of antibody neutralization will be facilitated by defining the structure of F and the structure of epitopes associated with neutralization. This will allow novel antigen design. Characterizing the chemistry and post-translational modifications in F will also promote improved vaccine antigen design. We have advanced our stabilized prefusion F protein into clinical evaluation in a Phase 1 clinical trial, VRC 317. In this trial, we are evaluating safety, tolerability, and immunogenicity of our stabilized prefusion F protein (DS-Cav1) with or with alum adjuvant in healthy adults. We have assessed the immune response to a single dose of DS-Cav1 in humans and found that DS-Cav1 vaccination elicits a 7-15-fold increase in neutralizing activity. Dissection of the antibody response showed that DS-Cav1 immunization elicited both antibodies that bind to the prefusion form of the protein and antibodies that bind to both the prefusion and postfusion conformation. The neutralizing activity elicited by DS-Cav1 vaccination is higher than that elicited by RSV infection, and 2-5 fold higher than neutralizing activity elicited by post-F or non-determined F structure. This work serves as a clinical proof-of-concept for structure-based vaccine design. In addition to neutralizing antibodies, the induction of both protective, disease-sparing CD8+ T cell responses are desirable in a vaccine candidate. These studies include evaluation of gene and vector-based vaccines expressing either the stabilized Pre-F protein or the wildtype F protein to vaccination with soluble Pre-F protein. We are also working with collaborators to assess immunogenicity of other vectors expressing our stabilized Pre-F protein.